Parasitic protozoans of the family Trypanosomatidae cause disease on a world wide scale in a variety of verterbrate, inverterbrate, and plant species. Trypanosoma cruzi, the etiologic agent of Chagas' disease, is a member of this family. Chagas disease is endemic throughout much of South and Central America afflicting over twenty million people. While the disease is essentially absent from North America, T. cruzi is indigenous to portions of the southwestern United States. T. cruzi has a complex life cycle involving obligatory developmental stages within the insect vector and the mammalian host. These stages include the non-dividing trypomastigotes (found in both the insect vector and the blood stream of the host) as well as the dividing epimastigotes (exclusively in the insect vector) and intracellular amastigotes. The only forms capable of invading vertebrate cells are the non-dividing trypomastigotes. An understanding of the genetic mechanisms involved in the control of the developmental cycle will undoubtedly add significantly to our understanding of the biology of this class of protozoan. To elucidate these genetic mechanisms we have chosen to study the expression of the polyubiquitin (PUB) and ubiquitin-fusion (FUS) genes of T. cruzi. These genes were chosen because many of the factors known to regulate their expression in other eukaryotic organisms are encountered by the parasite either as a consequence of their developmental life cycle or alterations in the growth conditions experienced during transmission. Our previous work has demonstrated that these genes are deferentially regulated in response to environmental stress or altered growth conditions. Further we have shown that at least two PUB genes are preceded by DNA sequences containing homologies to the heat shock elements found in other organisms. Before gene regulation can be understood at the molecular level, promoters and regulatory sequences must first be identified and characterized. Promoters have not been identified for any protein coding genes in trypanosomes therefore, we shall initially focus on the identification and characterization of the primary transcription products of the PUB and FUS genes. By mapping the 5' termini of the primary transcripts, regions containing potential promoters and/or regulatory regions will be identified. These regions will be further analyzed based on their protein binding characteristics.